Cloned (Comment) | Organism |
---|---|
expression of His-tagged wild-type and truncation mutant enzymes in Escherichia coli strain BL21(DE3) | Leishmania donovani |
Protein Variants | Comment | Organism |
---|---|---|
additional information | construction of 4 truncation enzyme mutants, i.e. mutants comprising residues 1-1058, residues 430-1058, residues 430-1236, or residues 430-785, deletion of 178 C-terminal residues remains without effect on the enzyme activity, while deletion of 451 and 429 residues from the C-terminus and the N-terminus, respectively, leads to complete loss of activity, the KCl requirement for mutant activity is altered, and the sensitivity to etoposide is eliminated, all mutants still bind to DNA, overview | Leishmania donovani |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
KCl | required for DNA cleavage, KCl is inhibitory above 400 mM | Leishmania donovani |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.33 | - |
ATP | recombinant 116 kDa mutant, ATP hydrolysis activity, in presence of DNA | Leishmania donovani | |
0.55 | - |
ATP | recombinant 116 kDa mutant, ATP hydrolysis activity, in absence of DNA | Leishmania donovani |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Ca2+ | divalent cation is required for ATP hydrolysis activity, optimal at 4 mM | Leishmania donovani | |
KCl | required for DNA cleavage, KCl is inhibitory above 400 mM | Leishmania donovani | |
Mg2+ | divalent cation is required for ATP hydrolysis activity, optimal at 6 mM | Leishmania donovani |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
40000 | - |
x * 116000, recombinant truncation mutant comprising residues 1-1058, SDS-PAGE, x * 70000, recombinant truncation mutant comprising residues 430-1058, SDS-PAGE, x * 89000, recombinant truncation mutant comprising residues 430-1236, SDS-PAGE, x * 40000, recombinant truncation mutant comprising residues 430-785, SDS-PAGE | Leishmania donovani |
70000 | - |
x * 116000, recombinant truncation mutant comprising residues 1-1058, SDS-PAGE, x * 70000, recombinant truncation mutant comprising residues 430-1058, SDS-PAGE, x * 89000, recombinant truncation mutant comprising residues 430-1236, SDS-PAGE, x * 40000, recombinant truncation mutant comprising residues 430-785, SDS-PAGE | Leishmania donovani |
89000 | - |
x * 116000, recombinant truncation mutant comprising residues 1-1058, SDS-PAGE, x * 70000, recombinant truncation mutant comprising residues 430-1058, SDS-PAGE, x * 89000, recombinant truncation mutant comprising residues 430-1236, SDS-PAGE, x * 40000, recombinant truncation mutant comprising residues 430-785, SDS-PAGE | Leishmania donovani |
116000 | - |
x * 116000, recombinant truncation mutant comprising residues 1-1058, SDS-PAGE, x * 70000, recombinant truncation mutant comprising residues 430-1058, SDS-PAGE, x * 89000, recombinant truncation mutant comprising residues 430-1236, SDS-PAGE, x * 40000, recombinant truncation mutant comprising residues 430-785, SDS-PAGE | Leishmania donovani |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Leishmania donovani | - |
- |
- |
Purification (Comment) | Organism |
---|---|
recombinant His-tagged wild-type and truncation mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography | Leishmania donovani |
Reaction | Comment | Organism | Reaction ID |
---|---|---|---|
ATP-dependent breakage, passage and rejoining of double-stranded DNA | active site mapping, catalytic residue Tyr775 in the gyrase half of the Leishmania enzyme is responsible for DNA binding, breakage, and rejoining | Leishmania donovani |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
kinetoplast DNA + ATP + H2O | cleavage of kinetoplast DNA | Leishmania donovani | ? | - |
? | |
additional information | ATPase activity is shown by the wild-type enzyme and the truncation mutant comprising residues 1-1058, the enzyme changes the DNA topology by coupling ATP hydrolysis to the transport of one DNA helix through a transient double-stranded break in another | Leishmania donovani | ? | - |
? | |
supercoiled pRYG DNA + ATP + H2O | relaxation of supercoiled pRYG DNA | Leishmania donovani | ? | - |
? |
Subunits | Comment | Organism |
---|---|---|
? | x * 116000, recombinant truncation mutant comprising residues 1-1058, SDS-PAGE, x * 70000, recombinant truncation mutant comprising residues 430-1058, SDS-PAGE, x * 89000, recombinant truncation mutant comprising residues 430-1236, SDS-PAGE, x * 40000, recombinant truncation mutant comprising residues 430-785, SDS-PAGE | Leishmania donovani |
dimer | multidomain homodimeric enzyme | Leishmania donovani |
Synonyms | Comment | Organism |
---|---|---|
Topoisomerase II | - |
Leishmania donovani |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
ATP | ATP stimulates dsDNA passage of the wild-type and the 116 kDa mutant enzyme | Leishmania donovani |